JPH0624967A - Substance having mutagen-suppressing action - Google Patents

Abstract

PURPOSE:To obtain a substance containing 3,4',5-trihydroxystilbene, having a simple structure and excellent mutagen-suppressing action. CONSTITUTION:An agent contains 0.2-0.4mg/ml 3,4',5-trihydroxystilbene. This substance exhibits strong suppressing action on induction of SOS reaction due to activated Trp-p-1. The concentration of the substance exhibits a positively proportional relationship to SOS reaction suppressive ratio and exhibits 72% SOS reaction suppressive ratio at 0.3mg/ml content and 98% SOS reaction suppressive ratio at 0.4mg/ml content. This substance can be expected as a carcinostatic substance.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a substance consisting of 3,4 ', 5-trihydroxystilbene which has a mutagen inhibiting effect.

[0002]

2. Description of the Related Art Conventionally, many studies have been carried out on a carcinogenic inhibitory substance consisting of an extract extracted from a plant,
As an extract from a monocotyledonous plant, a plant of Gramineae plant, Liliaceae plant, or Iridaceae plant origin is known. For example, in the origin of the Liliaceae plant, Cape Aloe leaves from South Africa (Japanese Patent Publication No. 46-28092), garlic or onions (Japanese Patent Laid-Open Nos. 48-98010 and 48).
-98011), which originates from the plant of the genus Tritonia, in the case of the origin of the iris family (JP-B-54
No. 28806) is known. In addition to this,
Ingredients extracted from medicinal carrot (JP-A-55-334)
56), an extract of Hiyodoribana belonging to the Asteraceae plant (Japanese Patent Laid-Open No. 55-45652), a component extracted from Achachatsuru (Japanese Patent Laid-Open No. 58-59921), and a component contained in Psycho (Japanese Laid-Open Patent Publication No. 62-187408) and other extracts containing saponins are known as substances having an anti-cancer effect.

As a stilbene derivative having an antitumor effect, 4,4'-bis (13.5-triazyl-6) is used.
-Amino) stilbene-2,2'-disulfonic acid derivative is known (Japanese Patent Publication No. 46-15503).

[0004]

However, the above-mentioned stilbene derivative has a very complicated structure, and its production is also difficult, and the appearance of a substance having a mutagen inhibitory effect that can be produced or extracted more easily has not been revealed. preferable. The present invention has been made in view of the above viewpoints, and an object thereof is to provide a substance having a simple structure and an excellent mutagen suppressing activity.

[0005]

The present inventors have completed the present invention as a result of various studies on substances having a mutagen suppressing action. That is, the mutagen inhibitor of the present invention is characterized by being composed of 3-4'-5-trihydroxystilbene (hereinafter, also simply referred to as trihydroxystilbene) shown in Chemical formula 1. Here, the term “consisting of trihydroxystilbene” is used to mean not only trihydroxystilbene itself but also the case of containing trihydroxystilbene (eg, solution, mixed powder containing other powders, etc.). It Usually, a solution as it is is used, or a solution prepared by dissolving powder or the like in a predetermined solvent immediately before use is used.

[0006]

[Chemical 1]

[0007]

EXAMPLES The present invention will be specifically described below with reference to examples. In this example, each sample (S) having the sample concentrations shown in Table 1 and the following was prepared, and addition of this sample caused induction of the SOS reaction by a certain amount of mutagen (activated T _rp -P-1). I investigated how much it was suppressed. This inhibitory effect was examined by a mutagen detection method (umu test) using the induction of SOS reaction as an index. Here, the "umu test" is a new type of mutagen detection test that utilizes the SOS reaction that is observed during DNA damage in Escherichia coli. This test method is described below, and has many advantages such as results in a short time. That is, 3 in LB medium (tryptone 1%, yeast extract 0.5%, salt 0.5%)
Test bacterial solution (test bacteria Salmonella ty
phimurium TA1535 / pSK1002) was inoculated into TGA medium (1% tryptone, 0.5% salt, 0.2% glucose with ampicillin added at a rate of 50 μg / ml) and shaken at 37 ° C. Cultured.

When the bacterial concentration reaches the logarithmic growth phase (A ₆₀₀ is 0.25 to 0.30), 2.0 ml of the bacterial solution is added.
Each of them was placed in a test tube, and 0.1 ml of a mutagen (activated T _rp -P-1 prepared as shown below), 0.1 ml of each sample, and 0.1 M phosphate buffer (pH 7.0) were added. )
0.3 ml was added and the mixture was cultured at 37 ° C. for 2 hours. After culturing, the bacterial solution was spun down to collect the cells, and then the cells were resuspended in physiological saline, the bacterial amount was measured in a part of this bacterial solution, and the β-galactosidase activity was measured in another part. Here, the logarithmic growth phase is the time when the number of bacteria and cells increases logarithmically, and is also called the exponential growth phase.

The β-galactosidase activity is measured by Mill
er's method (Miller, J. H: Experiments in molecular ge
netics, Coldspring Harbor Laboratory, New York,
P352-355 (1972)). That is, Z buffer 2.
After adding 0.25 ml of the above test bacterial solution to 25 ml,
50 μl of 1% aqueous solution of sodium dodecyl sulfate and 10 μl of chloroform were added and stirred vigorously. Substrate (o-nitro-phenyl-β-D-galactopyranoside 4 mg /
ml) 0.25 ml was added, and the mixture was reacted at 28 ° C. Then, after 15 minutes, 1.25 ml of 1 M Na ₂ CO ₃ was added to stop the reaction, and A ₄₂₀ , A ₅₅₀ and A were measured with a spectrophotometer.
₆₀₀ (absorbance) was measured.

Here, the β-galactosidase activity value is
It was calculated by the following formula. β-galactosidase activity value (unit) = 1000 (A ₄₂₀ −
1.75 × A ₅₅₀ ) /1.5×A ₆₀₀ The SOS reaction inhibition rate was calculated by the following formula. SOS reaction suppression rate (%) = [1- (AC) / (B-
C)] × 100 (%) where A is the β-galactosidase activity value when the sample (S) is added to the mutagen, and B is the β-galactosidase activity value induced only by the mutagen. , C shows the control β-galactosidase activity value, respectively.

The sample concentration is 10 mg / sample of the above sample (S).
Dissolved in pure water to a concentration of ml to make a stock solution, 1 mg (trihydroxystilbene) /2.5 m
1, 0.75mg / 2.5ml, 0.5mg / 2.5m
3 concentrations of 1 (ie 0.4 mg / ml, 0.3 m, respectively)
g / ml, 0.2 mg / ml), and samples of these 3 concentrations were investigated. The same amount of pure water was used as a control. Each test was conducted as one set of trials, and the average thereof was taken. Creation of activated T _rp -P-1
The T _rp -P-1 5mg dissolved in pure water 20 ml, and cultured at 37 ° C. by adding S9mix 20 ml thereto. After 20 minutes, 40 ml of acetone was added to stop the reaction, and the denatured protein was removed by centrifugation. This centrifugal supernatant was decompressed and concentrated (without acetone) by a rotary evaporator under ice cooling, and this concentrated liquid was _designated as activated T _rp -P-1. The activated T _rp -P-1 solution in the table is T _rp -P-
One concentration was 0.4 mg / ml. And
The activated T _rp -P-1 / trihydroxystilbene mixed solution had a trihydroxystilbene concentration shown in the table and 0.
4 shows a T _rp -P-1 concentration of 4 mg / ml. The results of the above tests are shown in Table 1.

Table 1 -------------------------------------------. Beta.-Galactosidase activity inhibition rate (%) −−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−− Control 176 ──Trihydroxystilbene solution (0.4 mg / ml) 168 _--Activated Trp-P-1 solution (0.4 mg / ml) 423 ---------------------------------. -------- activated T _rp -P-1 / Torihito Bu Rokishisuchiruben mixture 0.4 mg / ml 181 98.0 Torihito Bu Rokishisuchiruben concentration 0.3mg / ml 245 72.1 0.2mg / ml 280 57.9 −−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−

As shown in Table 1 above, the concentration of trihydroxystilbene is directly proportional to the SOS reaction inhibition rate, and particularly when the concentration is 0.4 mg / ml,
The suppression rate improved to 98.0%. This causes trihydroxystilbene to undergo S by activation T _rp -P-1.
It is clear that it has a strong inhibitory effect on the induction of the OS response. The present invention is not limited to the specific examples described above, and various modifications may be made within the scope of the present invention depending on the purpose and application. That is, the concentration of trihydroxystilbene may exceed 0.4 mg / ml, and the solvent for forming the solution is not limited to water,
Other solvents or mixed solvents may be used depending on the purpose. Further, various additives may be added to the use solution.

[0014]

INDUSTRIAL APPLICABILITY As described above, the substance containing 3-4'-5-trihydroxystilbene has a simple structure and an excellent suppressive effect of suppressing mutagen. Therefore, it can be sufficiently expected as a carcinogen. When the trihydroxystilbene concentration is 0.3 mg / ml or more, the SOS reaction inhibition rate is 70% or more, which is an extremely excellent effect.

Claims (2)

[Claims] 1. A substance having a mutagen inhibitory action, which comprises 3,4 ′, 5-trihydroxystilbene. 2. The substance having a mutagen inhibiting activity according to claim 1, wherein the concentration of the 3,4 ′, 5-trihydroxystilbene is 0.3 mg / ml or more.